IntroductionImmune checkpoint inhibitors (ICIs) targeting the PD-1/PD-L1 axis have expanded treatment options for hepatocellular carcinoma (HCC), yet only a subset of patients achieves durable responses. Limited efficacy reflects gaps in understanding PD-L1 regulation, including its biochemical heterogeneity, subtype-specific stability, and effects on tumor–immune interactions. We aimed to dissect these mechanisms and identify integrated biomarkers predictive of immunotherapy response.MethodsHuman HCC and paired non-tumoral tissues, an HBV-driven transgenic mouse model, and five HCC cell lines representing Hoshida/Caruso subtypes were analyzed. PD-L1 expression, glycosylation, and stability were assessed by Western blot, flow cytometry, and immunohistochemistry. Functional assays included co-cultures of HCC cells with primary activated PBMCs from multiple healthy donors. Cells were treated with durvalumab or atezolizumab, in the presence or absence of IFN-γ stimulation. PD-L1 turnover was evaluated using cycloheximide and proteasome inhibition.ResultsPD-L1 protein was increased in ~60% of tumors, particularly in virally driven HCC. Elevated PD-L1 in non-tumoral liver was associated with higher recurrence risk. Biochemical profiling revealed multiple PD-L1 species: mature N-glycosylated and intermediate forms indicative of enhanced stability were enriched in tumors. Glycosylated PD-L1 displayed prolonged half-life in S1/CL3 subtypes, whereas S2/CL1 cells exhibited rapid, proteasome-dependent turnover. PD-L1 abundance alone did not predict immune susceptibility: S1-like cells, despite higher PD-L1, were highly sensitive to CD8+ T-cell–mediated killing and PD-L1 blockade, whereas S2-like cells were more resistant to cytotoxicity. ICIs induced donor-dependent cytotoxicity, with variable responder profiles; durvalumab outperformed atezolizumab, consistent with reduced glycosylation dependence. IFN-γ priming enhanced PD-L1 expression and restored immune responsiveness in low-responder co-cultures. Overall, tumors with stable glycosylated PD-L1 (S1-like) rely on PD-L1–mediated immune suppression and may benefit from PD-L1–targeted therapies, whereas tumors with low or rapidly turned-over PD-L1 (S2-like) exhibit dynamic PD-L1 regulation with increased reliance on de novo synthesis, limiting responsiveness to PD-L1 blockade alone and supporting the need for combinatorial approaches.ConclusionsPD-L1 regulation in HCC depends on molecular subtype, post-translational stability, and host immune competence. Integrating these factors provides a framework for guiding patient selection and optimizing immunotherapy.
Background/Objectives: Metabolic dysfunction-associated steatotic liver disease (MASLD) is a common chronic liver disorder linked to obesity, insulin resistance, and dyslipidemia. Nutrition plays a central role in modulating hepatic lipid metabolism, oxidative stress, and inflammation, yet practical, evidence-based dietary strategies remain limited. This study aimed to develop Mediterranean diet-based meal plans with varying macronutrient compositions and to characterize their nutritional profiles, as well as to evaluate them using established nutritional indices and diet score calculations, such as the Dietary Inflammatory Index, Dietary Antioxidant Index, and dietary lipid indices. Methods: Clinical practice guidelines (CPGs) from various academic and professional organizations were reviewed to assess current non-pharmacological treatments for MASLD, with a focus on determining whether the Mediterranean diet is the most recommended dietary pattern. Traditional, low-carbohydrate, and low-fat MedDiet patterns were translated into food-based meal plans. A 7-day meal plan was developed and analyzed for nutrient composition, then evaluated using the Dietary Inflammatory Index (DII), Dietary Antioxidant Index (DAI), Dietary Lipophilic Index (DLI), and Dietary Lipophilic Load (DLL). A Western diet (WD) that is characterized by ultra-processed food (UPF) was included as a comparative reference. Results: The validated dietary score calculations showed that all MedDiet patterns demonstrated consistently low DII scores (-2.00 to -2.81) and high DAI scores (3 to 20.03), whereas the WD showed high DII scores (5.0 to 6.09) and low DAI scores (-12.47 to -17.99). Despite these variations in macronutrients, the menu developed in the study on three MedDiet patterns showed negative DII and positive DAI scores. When comparing the traditional MedDiet with the WD, which have similar macronutrient distributions, the WD was characterized by less favorable DII and DAI scores. Conclusions: This study provides a descriptive, guideline-informed framework for Mediterranean diet-based meal plans with varying macronutrient compositions. Utilizing DII, DAI, DLI, and DLL offers a potential framework for designing dietary interventions. Further validation through clinical studies is needed to justify the potential for practical and digital translation. Nevertheless, the study provides initial insights that may inform future research on nutritional approaches for MASLD integrating dietary indices.
Aim: This study characterized pre-packaged foods in Southeast Asia using Nutri-Score, NOVA, and Multiple Traffic Light systems; cross-classified products by Nutri-Score and NOVA; and assessed their ability to discriminate products by nutrient content. Methods: Data were obtained from Open Food Facts in October 2024. Of 29,789 products identified in Southeast Asia, duplicates and products with incomplete information were excluded, resulting in 4,668 pre-packaged food products from eight Southeast Asian countries. Salt, sugar, total fat, and saturated fat contents were evaluated using the Multiple Traffic Light system. Nutri-Score data were available for 4,565 products, while 2,068 products had complete information for both Nutri-Score and NOVA and were included in cross-classification analysis. Spearman correlation analyses examined relationships among Nutri-Score, NOVA, and nutrient profiles. Results: Most products were of lower nutritional quality, with 52% classified as Nutri-Score D or E and 71% categorized as ultra-processed (NOVA 4). Multiple Traffic Light classification showed that beverages and sweet products had the highest proportion high in sugar, while beverages, cereals, and dish meals were most frequently high in salt. Sweets and snacks were the main contributors to products high in total fat and saturated fat. Cross-classification showed that as Nutri-Score decreased from A to E, the prevalence of ultra-processed foods consistently increased, while NOVA 1 became nearly negligible, indicating a trend between poorer nutritional quality and higher processing. Nutri-Score showed weak-to-moderate positive correlations with sugar (r = 0.355, p < 0.001), saturated fat (r = 0.364, p < 0.001), and salt (r = 0.190, p < 0.001). NOVA showed weaker but significant correlations with sugar (r = 0.149, p < 0.001) and salt (r = 0.138, p < 0.001). Conclusions: Nutri-Score was associated with nutrient-rich foods, while NOVA highlighted processing levels. Combined use offers a more comprehensive evaluation than either system alone. Integrating nutrient profiling and processing classification in front-of-pack labeling may guide consumer choices and public health efforts to reduce diet-related NCDs in Southeast Asia.
Chronic low-grade inflammation is a key feature of obesity-associated dysglycemia, yet substantial heterogeneity exists in inflammatory responses among individuals with normoglycemia, prediabetes, and type 2 diabetes mellitus (T2DM). Whether circulating inflammatory protein profiles define distinct patient phenotypes beyond conventional glycemic classification remains incompletely understood. In this cross-sectional analysis of 142 individuals with severe obesity, plasma inflammatory proteins were quantified using Olink proximity extension assay technology. Subjects were stratified by glycemic status (noDM, normoglycemia; PreDM, prediabetes and T2DM) while maintaining comparable distributions of metabolic dysfunction-associated steatotic liver disease. Differential expression analyses were performed across glycemic groups, and unsupervised topological data analysis (TDA) was applied to identify inflammatory protein-based patient subgroups. Several inflammatory proteins were significantly upregulated in T2DM and PreDM compared with noDM, with interleukin-8 (IL-8), Fms-relatedlike tyrosine kinase 3 ligand (Flt3L), and CUB domain containing protein (CDCP1) showing the largest significant differences. NPX distributions of these proteins exhibited gradual increases across glycemic stages with substantial inter-individual variability. TDA identified seven clusters defined by distinct inflammatory protein signatures. One cluster was enriched for individuals with T2DM and characterized by coordinated upregulation of IL-8, Flt3L, CDCP1, and additional immune- and cytokine-related proteins, whereas other clusters displayed alternative inflammatory profiles that were not explained by glycemic status alone. Inflammatory proteomic profiling in severe obesity reveals both glycemia-associated protein changes and distinct inflammatory phenotypes that transcend conventional clinical classification. Integration of differential expression analysis with TDA highlights heterogeneity in inflammatory states, supporting a hypothesis-generating framework for future studies aimed at validating these proteomic patterns and clarifying their longitudinal relevance in obesity-related dysglycemia.
Ultra-processed foods (UPFs) are increasingly consumed worldwide, and higher consumption has been linked to higher risk of non-communicable diseases (NCDs). This study examined the nutritional composition and quality, energy density, and hyperpalatability of UPFs available in the Italian food market according to their literature-derived NCD-risk categories. Products were classified into main food groups and subgroups and further categorized according to reported associations with NCD risk: positive, negative, or no reported association. Differences between the main food groups were analyzed in RStudio using non-parametric tests, with results summarized using medians and interquartile ranges. A total of 13,025 UPF products were included in the analysis. UPFs showed substantial variation in nutritional composition, nutritional quality, nutrients-of-concern levels, energy density, and hyperpalatability across the main food groups and literature-derived NCD-risk categories. Snacks represented the largest UPF group and showed the most consistently unfavorable profile, including high median energy, carbohydrate, saturated fat, and sugar contents, a high proportion of Nutri-Score D and E products, and the highest proportion of medium- and high-energy-density products. Overall, 64.3% of UPFs were classified as Nutri-Score D or E, indicating generally lower nutritional quality. However, unfavorable nutritional characteristics were not limited to UPF subgroups reportedly associated with increased NCD risk; some subgroups reportedly associated with lower or no reported NCD risk also contained products with poor nutritional quality, high energy density, or hyperpalatable profiles. These findings suggest that UPFs are not nutritionally uniform and that assessment of UPF-related health relevance should consider food group, subgroup, nutrient profile, energy density, and hyperpalatability in addition to processing classification.
